Article Figures & Data
Figures
- FIG 1
Structures of Gwt1 inhibitors.
- FIG 2
Efficacy of APX001 in a murine model of coccidioidomycosis. Mice were infected intranasally with C. immitis RS arthroconidia and 50 mg/kg of APX001 was administered twice daily for 5 days beginning 7 days postinfection. Mice were sacrificed on day 13, 1 day after the last day of treatment, and colony counts were assessed from the lung and spleen. Each symbol represents one mouse. The horizontal lines show the geometric mean and SEM for the lung and spleen colony counts (CFU). Horizontal lines in the weight panel correspond to the calculated mean weight. The differences in mean weight of treated and control mice on days 11 and 13 were analyzed by two-way ANOVA (GraphPad Prism) and were highly significant (P < 0.001).
- FIG 3
ABT alone has no antifungal effect in mice. Infected mice were treated with a single daily dose of ABT for 5 days and sacrificed on day 13, the day after the last ABT dose. Fungal colony counts were log transformed. Geometric mean CFU/organ values ± the SEM were calculated and compared using an unpaired t test (Prism, v7.01). The mean weights ± the SEM were calculated, and there was no significant difference in the weights of untreated and ABT-treated mice on day 15 after infection.
- FIG 4
Reduction in fungal burden upon treatment with three Gwt1 prodrugs in a mouse model of pulmonary coccidioidomycosis. Mice were infected and treated as in Fig. 2, except that mice were pretreated with 50 mg/kg ABT by oral gavage 2 h prior to the administration of APX prodrugs or buffer starting 7 days after infection. Mice were weighed at the start and conclusion of the experiment and were sacrificed 1 day after their last dose. After log10 transformation, values for the geometric mean CFU/organ ± the SEM were calculated and compared using an unpaired t test (Prism, v7.01). If there were >2 groups, the differences in the means of treated and control groups were compared using Dunnett’s ANOVA test. A P value of ≤0.05 was considered statistically significant.
- FIG 5
Reduction in fungal burden upon treatment with APX001 and APX2097 in comparison to fluconazole. Mice were infected and treated with the ABT and APX prodrugs as in Fig. 3. Fluconazole was administered orally twice daily. Values for the geometric mean CFU/organ ± the SEM were calculated and compared using a paired t test (Prism, v7.01). If there were >2 groups, the differences in the means of treated and control groups were compared using Dunnett’s ANOVA test. All of the treatment groups had significant lower colony counts than the untreated control in lungs and spleen (P < 0.001). Only the untreated mice had a statistically significant eight loss on day 13 after infection compared to their starting weight (P = 0.006).
- FIG 6
Histological analysis of lung tissue sections in control versus APX001- and APX2097-treated mice. Mice were infected with C. immitis RS as described in Materials and Methods and then treated once daily with 50 mg/kg ABT plus APX001 or APX2097 at 26 mg/kg for 5 days. Control mice received only ABT. Lungs were removed a few hours after the last dose, fixed in glutaraldehyde, and then stained with PAS prior to microscopic examination (×20 magnification). (A) The control lungs showed many spherules in all stages of development and a myriad of endospores from ruptured spherules, surrounded by acute and chronic inflammatory cells. (B) APX001-treated mice had many small, immature spherules that were primarily inside macrophages. There were no fully grown spherules and few if any endospores. (C) The lungs of APX2097-treated mice had an appearance similar to the lungs of APX001-treated mice.
- FIG 7
Comparison of Kaplan-Meir survival curves and end-of-treatment weights of mice treated with APX001, APX2097, or fluconazole compared to untreated controls. Mice were infected and treated as described in Fig. 3. The arrows show the days of treatment. Kaplan-Meir survival curves were compared by log rank analysis (Prism, v7.01). All three treatment groups survived significantly longer than the control mice. Differences between the three treatment groups was also significant. The mean body weights of the three treatment groups and the untreated control on day 14 posttreatment were compared by ANOVA (Tukey’s multiple-comparison test; Prism, v7.01). There were no significant differences in the weights of fluconazole-, APX001-, and APX2097-treated mice. *, P < 0.01 for both graphs.
Tables
- TABLE 1
In vitro susceptibility profiles
Strain MEC (μg/ml), APX001A MIC (μg/ml)a APX001A FLC AMB POS C. immitis RS 0.002–0.004 8 >16 0.125 0.06–0.125 C. posadasii C735 0.004 0.03 >16 0.25 0.06–0.125 C. posadasii Silvera 0.008 8 >16 0.25 0.03 ↵a The MIC value was read at 100% inhibition. FLC, fluconazole; AMB, amphotericin B; POS, posaconazole.
- TABLE 2
Activity of Gwt1 inhibitors versus C. immitis and C. posadasii
Strain or parameter Source MEC (μg/ml) MIC (μg/ml) APX001A APX2020 APX2041 POS C. immitis RS Lab 0.002–0.004a 0.002–0.004 0.002–0.004 0.06–0.125 B2358 CDC 0.004 0.004 0.000125 0.016 F40 Clinical 0.004 0.002 0.001 0.125 F1 Clinical 0.002 0.001 0.001 0.125 UCSD2 Clinical 0.001 0.001 0.00025 0.125 C. posadasii F6 Clinical 0.016 0.004 0.001 0.125 Silvera Lab 0.008 0.008 0.004 0.03 F5 Clinical 0.008 0.004 0.001 0.016 C735 Lab 0.004 0.002 0.002 0.06–0.125 D2A Clinical 0.004 0.002 0.001 0.03 Parameters GM 0.004 0.002 0.001 0.054 MEC90/MIC90 0.008 0.004 0.002 0.125 ↵a The lower value of the susceptibility range was used to calculate the geometric mean (GM) and 90% minimal effective concentration (MEC90).
